Encapsulation of hemoglobin in phospholipid liposomes: characterization and stability

Abstract

Hb is encapsulated in liposomes of different lipid composition. The resulting dispersion consists primarily of multilamellar liposomes (hemosomes) of a wide particle size distribution (diameter ranging mainly between 0.1 and 1 .mu.m). The encapsulation efficiency is significantly larger with liposomes containing negatively charged lipids as compared to liposomes made of phosphatidylcholine. The integrity of the phospholipid bilayer is maintained in the presence of Hb. The reaction rate of CO binding to encapsulated Hb is reduced compared to that of free Hb, but it is still greater than that observed in [human] red blood cells. Hb encapsulated in liposomes made from negatively charged phospholipids is less stable than Hb entrapped in isoelectric phosphatidylcholine. The instability of Hb is due to the protein interacting with the negatively charged lipid bilayer. This interaction leads in turn to Hb denaturation, possibly involving the dissociation of the heme group from the heme-globin complex. The nature of the negatively charged phospholipid is important in promoting the interaction with Hb, the effect being in the order phosphatidic acid > phosphatidylinositol .simeq. phosphatidylglycerol > phosphatidylserine. The presence of equimolar amounts of cholesterol in the phospholipid bilayer has a stabilizing effect on Hb. This effect is pronounced with saturated phospholipids, but it is also observed, though to a lesser extent, with unsaturated ones, indicating that the bilayer fluidity has a modulating effect. The presence of cholesterol possibly interferes with secondary interactions following the binding of Hb to the negatively charged lipid bilayer.